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Ectopic Expression of Mitochondria Endonuclease Pnu1p from Schizosaccharomyces pombe Induces Cell Death of the Yeast

  • Oda, Kaoru (Department of Biochemistry, Faculty of Science, Okayama University of Science) ;
  • Kawasaki, Nami (Department of Biochemistry, Faculty of Science, Okayama University of Science) ;
  • Fukuyama, Masashi (Department of Biochemistry, Faculty of Science, Okayama University of Science) ;
  • Ikeda, Shogo (Department of Biochemistry, Faculty of Science, Okayama University of Science)
  • Published : 2007.11.30

Abstract

Endonuclease G (EndoG) is a mitochondrial non-specific nuclease that is highly conserved among the eukaryotes. Although the precise role of EndoG in mitochondria is not yet known, the enzyme is released from the mitochondria and digests nuclear DNA during apoptosis in mammalian cells. Schizosaccharomyces pombe has an EndoG homolog Pnu1p (previously named SpNuc1) that is produced as a precursor protein with a mitochondrial targeting sequence. During the sorting into mitochondria the signal sequence is cleaved to yield the functionally active endonuclease. From the analogy to EndoG, active extramitochondrial Pnu1p may trigger cell killing by degrading nuclear DNA. Here, we tested this possibility by expressing a truncated Pnu1p lacking the signal sequence in the extramitochondrial region of pnu1-deleted cells. The truncated Pnu1p was localized in the cytosol and nuclei of yeast cells. And ectopic expression of active Pnu1p led to cell death with fragmentation of nuclear DNA. This suggests that the Pnu1p is possibly involved in a certain type of yeast cell death via DNA fragmentation. Although expression of human Bak in S. pombe was lethal, Pnu1p nuclease is not necessary for hBak-induced cell death.

Keywords

References

  1. Arnoult, D., Gaume, B., Karbowski, M., Sharpe, J. C., Cecconi, F. and Youle, R. J. (2003) Mitochondrial release of AIF and EndoG requires caspase activation downstream of Bax/Bak-mediated permeabilization. EMBO J. 22, 4385-4399. https://doi.org/10.1093/emboj/cdg423
  2. Craven, R. A., Griffiths, D. J., Sheldrick, K. S., Randall, R. E., Hagan, I. M. and Carr, A. M. (1998) Vectors for the expression of tagged proteins in Schizosaccharomyces pombe. Gene 221, 59-68. https://doi.org/10.1016/S0378-1119(98)00434-X
  3. Endoh, H. and Kobayashi, T. (2006) Death harmony played by nucleus and mitochondria: nuclear apoptosis during conjugation of tetrahymena. Autophagy 2, 129-131. https://doi.org/10.4161/auto.2.2.2368
  4. Huang, K. J., Ku, C. C. and Lehman, I. R. (2006) Endonuclease G: a role for the enzyme in recombination and cellular proliferation. Proc. Natl. Acad. Sci. USA 103, 8995-9000. https://doi.org/10.1073/pnas.0603445103
  5. Ikeda, S. and Kawasaki, N. (2001) Isolation and characterization of the Schizosaccharomyces pombe cDNA encoding the mitochondrial endonuclease. Biochim. Biophys. Acta 1519, 111-116. https://doi.org/10.1016/S0167-4781(01)00206-8
  6. Ink, B., Zornig, M., Baum, B., Hajibagheri, N., James, C., Chittenden, T. and Evan, G. (1997) Human Bak induces cell death in Schizosaccharomyces pombe with morphological changes similar to those with apoptosis in mammalian cells. Mol. Cell. Biol. 17, 2468-2474. https://doi.org/10.1128/MCB.17.5.2468
  7. Irvine, R. A., Adachi, N., Shibata, D. K., Cassell, G. D., Yu, K., Karanjawala, Z. E., Hsieh, C. L. and Lieber, M. R. (2005) Generation and characterization of endonuclease G null mice. Mol. Cell. Biol. 25, 294-302. https://doi.org/10.1128/MCB.25.1.294-302.2005
  8. Jurgensmeier, J. M., Krajewski, S., Armstrong, R. C., Wilson, G. M., Oltersdorf, T., Fritz, L. C., Reed, J. C. and Ottilie, S. (1997) Baxand Bak-induced cell death in the fission yeast Schizosaccharomyces pombe. Mol. Biol. Cell 8, 325-339. https://doi.org/10.1091/mbc.8.2.325
  9. Kobayashi, T. and Endoh, H. (2005) A possible role of mitochondria in the apoptotic-like programmed nuclear death of Tetrahymena thermophila. FEBS J. 272, 5378-5387. https://doi.org/10.1111/j.1742-4658.2005.04936.x
  10. Li, L. Y., Luo, X. and Wang, X. (2001) Endonuclease G is an apoptotic DNase when released from mitochondria. Nature 412, 95-99. https://doi.org/10.1038/35083620
  11. Low, C. P., Liew, L. P., Pervaiz, S. and Yang, H. (2005) Apoptosis and lipoapoptosis in the fission yeast Schizosaccharomyces pombe. FEMS Yeast Res. 5, 1199-1206. https://doi.org/10.1016/j.femsyr.2005.07.004
  12. Low, R. L. (2003) Mitochondrial Endonuclease G function in apoptosis and mtDNA metabolism: a historical perspective. Mitochondrion 2, 225-236. https://doi.org/10.1016/S1567-7249(02)00104-6
  13. Madeo, F., Herker, E., Wissing, S., Jungwirth, H., Eisenberg, T. and Frohlich, K. U. (2004) Apoptosis in yeast. Curr. Opin. Microbiol. 7, 655-660. https://doi.org/10.1016/j.mib.2004.10.012
  14. Nakashima, A., Hasegawa, T., Mori, S., Ueno, M., Tanaka, S., Ushimaru, T., Sato, S. and Uritani, M. (2006) A starvationspecific serine protease gene, isp6+, is involved in both autophagy and sexual development in Schizosaccharomyces pombe. Curr. Genet. 49, 403-413. https://doi.org/10.1007/s00294-006-0067-0
  15. Nakashima, A., Yoshida, M., Nakayama, K., Kato-Furuno, A., Ueno, M., Ushimaru, T. and Uritani, M. (2002) Genes for a nuclease and a protease are involved in the drastic decrease in cellular RNA amount in fission yeast cells during nitrogen starvation. J. Biochem. (Tokyo) 131, 391-398. https://doi.org/10.1093/oxfordjournals.jbchem.a003114
  16. Ohsato, T., Ishihara, N., Muta, T., Umeda, S., Ikeda, S., Mihara, K., Hamasaki, N. and Kang, D. (2002) Mammalian mitochondrial endonuclease G. Digestion of R-loops and localization in intermembrane space. Eur. J. Biochem. 269, 5765-5770. https://doi.org/10.1046/j.1432-1033.2002.03238.x
  17. Parrish, J., Li, L., Klotz, K., Ledwich, D., Wang, X. and Xue, D. (2001) Mitochondrial endonuclease G is important for apoptosis in C. elegans. Nature 412, 90-94. https://doi.org/10.1038/35083608
  18. Sakem, B. and Kohli, J. (2007) The end1 gene of Schizosaccharomyces pombe coding for a DNase is identical with the pnu1 gene coding for an RNase. Yeast 24,11-16. https://doi.org/10.1002/yea.1427
  19. Schafer, P., Scholz, S. R., Gimadutdinow, O., Cymerman, I. A., Bujnicki, J. M., Ruiz-Carrillo, A., Pingoud, A. and Meiss, G. (2004) Structural and functional characterization of mitochondrial EndoG, a sugar non-specific nuclease which plays an important role during apoptosis. J. Mol. Biol. 338, 217-228. https://doi.org/10.1016/j.jmb.2004.02.069
  20. Torgler, C. N., de Tiani, M., Raven, T., Aubry, J. P., Brown, R. and Meldrum, E. (1997) Expression of bak in S. pombe results in a lethality mediated through interaction with the calnexin homologue Cnx1. Cell Death Differ. 4, 263-271. https://doi.org/10.1038/sj.cdd.4400239
  21. van Loo, G., Schotte, P., van Gurp, M., Demol, H., Hoorelbeke, B., Gevaert, K., Rodriguez, I., Ruiz-Carrillo, A., Vandekerckhove, J., Declercq, W., Beyaert, R. and Vandenabeele, P. (2001) Endonuclease G: a mitochondrial protein released in apoptosis and involved in caspase-independent DNA degradation. Cell Death Differ. 8, 1136-1142. https://doi.org/10.1038/sj.cdd.4400944
  22. Zhang, Q., Chieu, H. K., Low, C. P., Zhang, S., Heng, C. K. and Yang, H. (2003) Schizosaccharomyces pombe cells deficient in triacylglycerols synthesis undergo apoptosis upon entry into the stationary phase. J. Biol. Chem. 278, 47145-47155. https://doi.org/10.1074/jbc.M306998200

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